Enzymes associated with glycogen metabolism and glycerol synthesis in larvae of the Shonai ecotype of the rice stem borer, Chilo suppressalis, were investigated over the winter in 2000-2001. Glycerol content was scarcely detected in September and October, rapidly increased in November and December, peaked in January, and then decreased. Glycogen was converted to glycerol over the winter until February, and glycerol was reconverted to glycogen in March. The trehalose content remained constant over the winter. The activities of enzymes associated with glycerol synthesis changed with the season. Glycerol accumulation was accomplished by activation of glycogen phosphorylase, inhibition of fructose-1,6-bisphosphatase and pyruvate kinase, and activation of enzymes associated with glycerol synthesis, mainly glyceraldehyde-3-phosphatase and polyol dehydrogenase with glyceraldehyde activity. These changes led to a diversion of triose phosphates into the pathway of glycerol synthesis. However, activities of the two initial enzymes of the hexose monophosphate shunt were not activated and remained relatively constant, but high during the period of active glycerol synthesis. Both decreasing temperature in the field and the transition from the diapause to the post-diapause state may be responsible for the changes in activities of enzymes associated with glycerol synthesis.